Tubulin and Microtubules

Abstract

Microtubules are found in almost all types of eukaryotic cell. Their principal constituents are the highly conserved proteins
of the tubulin family, in the form of longitudinal protofilaments of tubulin heterodimers. The conserved lateral interactions
produce a characteristic 2D lattice. Tubulin structure and conformational changes are now known to atomic resolution. Its
GTP‐dependent dynamic self‐assembly and disassembly is modulated by a host of other proteins. This dynamic activity and a
more passive role as tracks for interaction with motor proteins, kinesin and dynein, allow microtubules to perform vital roles
in various forms of cell motility. They are an essential component of cell division, provide oriented tracks for the transport
of cellular organelles and vesicles and are responsible for the relative positioning of cellular compartments. Thus, they
are important targets in the control of cancer cells, as their essential activity is severely affected by a variety of drugs.

Key Concepts

The assembled sheet has a slightly variable curvature, producing microtubules, with around 13 protofilaments but not strictly
defined unless they are grown from a γ‐tubulin template or given a curvature appropriate to exactly 13 protofilaments by accessory
proteins

The structure of microtubules has been solved to near‐atomic level both with GTP homologues bound to β‐tubulin and with GDP
bound; also a depolymerised protofilament structure in a longitudinally curved conformation is known

Hydrolysis of bound GTP enables microtubules to assemble with ‘dynamic instability’, behaviour that depends on stochastic
disassembly when the GTP ‘cap’ is lost from a growing end

Microtubule behaviour is modified by many different accessory proteins that may act just at the tips or everywhere on the
lattice wall

To play different roles according to their positions in cells, the properties of microtubules may vary owing to different
tubulin isotypes and be modified by a wide range of associated proteins

Changes can also vary with time (‘ageing’) as a result of post‐translational modification of tubulin and of some accessory
proteins

Microtubules are employed in axonemes of cilia and flagella, where the motor protein dynein interacts with them to produce
beating while other linking components help control the wave‐form

Cytoplasmic microtubules provide tracks for vesicles and other cargoes driven by kinesins and cytoplasmic dynein

Mitotic and meiotic spindles are assembled from microtubules, which interact with kinetochores and with multiple motor proteins

Microtubules are susceptible to many small molecules that depolymerise or hyperstabilise them and thus serve as anti‐cancer
drugs.

Figure 1. (a,b) Microtubules in cells viewed by immunofluorescence light microscopy. DNA is labelled in blue, all microtubules in green,
and glutamylated tubulin in red. The central spindle of the cell in anaphase (a) and the midbody microtubules between cells
in the final stage of division (b) appear yellow (green plus red), where the tubulin is more modified. (c) A single microtubule
as seen by cryo‐electron microscopy. (a,b) Courtesy of Dr Carsten Janke. (c) Taken by Dr Keiko Hirose.

Figure 2. Structure of microtubules. (a) Atomic structure of the αβ‐tubulin heterodimer (PDB code 1JFF) shown as a ribbon diagram; GTPase
domains in red, intermediate/activation domains in blue,
‐terminal/outer‐surface domains in green, and central helices in yellow. (b) Assembly of heterodimers into longitudinal protofilaments,
which in turn associate into sheets and microtubules. Side‐to‐side association of the protofilaments in two possible ways
can produce an A‐lattice (symmetrical for the 13‐protofilament microtubule shown) or a B‐lattice (with an A‐lattice‐like seam
for 13 or 14 protofilaments, though not for 15 or 16). (c) Typical features of the two ends of a microtubule. The minus ends
cells are mostly capped by a γ‐tubulin ring complex that provides a template for the growth of 13 protofilaments. The plus
ends often remain free to grow by addition of heterodimers to sites with GTP bound on the exposed β‐tubulin and shrink by
loss of subunits that have caused GTP‐hydrolysis.

Figure 3. Structural changes in protofilaments. (a) αβ‐tubulin heterodimers as viewed from inside a microtubule. The two globular domains
in each subunit are coloured red (GTPase) and blue; GDP, bound to the GTPase domains, is shown as space‐filling atoms (grey).
The globular domains are separated by the core helix (yellow); Taxol binds in an adjacent pocket, where it promotes the straight
conformation. The M‐loops make lateral contact with loops in the GTPase domains of another protofilament. At an active interface
within a protofiilament, the T7 loop of α‐tubulin triggers GTP hydrolysis between heterodimers. The spacing there is greater
when GTP is present than it is after hydrolysis, when loops around the nucleotide change conformation. The passive intra‐dimer
interface does not change. (b) Stathmin (in grey) binds to two tubulin heterodimers in a curved conformation, in which there
is bending at the passive and active interfaces. The binding sites are shown of some drugs (colchicine and vinblastine) that
promote curvature and hence depolymerisation. Stathmin sequesters heterodimers to reduce the concentration of tubulin available
for assembly.

Figure 4. Diagrams showing how the orientations of microtubules vary during the cell cycle. (a) A typical motile cell, such as a fibroblast,
in interphase. Most microtubules grow out towards the cell periphery keeping their minus ends associated with the centrosomal
region. This structure contains a pair of centrioles surrounded by a cloud of proteins that include γ‐tubulin. The plus ends
grow out dynamically, with specific proteins tracking the microtubule tips. These may interact eventually with specific membrane‐associated
proteins or with the cortical layer of actin filaments (yellow). See also: Cell Locomotion (b) Vertebrate mitotic spindle depicted in metaphase (chromosomes paired at the mid‐plane of the spindle) and in anaphase
(chromosomes moving towards the poles). The centrosome at each pole, containing one old and one new centriole, acts as a focus
for microtubule minus ends.

Figure 5. (a) Cross section through a typical axoneme, viewed from the tip, that is, from the plus ends of the microtubules. Microtubules
were first seen in flagellar and ciliary axonemes. (b) Enlarged cross section of one of the doublet microtubules. Nine doublet
microtubules and a central core structure are linked together by a complex variety of accessory molecules. Centrioles (Figure) are built from nine triplet microtubules, with a C‐microtubule attached to each B‐microtubule; their A tubules do not have
dynein arms attached.

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